Solid-phase peptide synthesis spectrometry

S. Beranova-Giorgianni and D. M. Desiderio. Fast Atom Bombardment Mass Spectrometry of Synthetic Peptides. In Methods in Enzymology Solid-Phase Peptide Synthesis, ed. G. B. Fields. Methods in Enzymology 289. Academic Press, San Diego, 1997, 478-499. 

Maux, D. Enjalbal, C. Martinez, J. Aubagnac, J.-L. Combarieu, R. Static Secondary Ion Mass Spectrometry to Monitor Solid-Phase Peptide Synthesis. J. Am. Soc. Mass Spectrom. 2001, 72, 1099-1105. 

Quenched fluorescent substrates (QFS). The QFS listed in Tables 1 and 2 were synthesized by standard solid phase peptide synthesis techniques, and purified by HPLC (see Notes 1 and 2). The identity of each QFS was verified by mass spectrometry. The BK-based QFS (compound no. 26, Table 2), published previously by others (1), was solubiUzed in DMSOiethanol (50 50) at a stock concentration of 2 mg/mL (=1.2 mM), and kept at -20°C (see Note 3). Stock QFS was diluted 1 10 in assay buffer just prior to use. 

Solid phase peptide synthesis of the c-Myc and the Max LZs, characterization by mass spectrometry, purification by reversed-phase HPLC and the formation of the disulfide linked c-Myc-Max heterodimeric LZ have been described elsewhere (19). 

K McMellop, W Davidson, G Hansen, D Freeman, N Pallai. The characterization of crude products from solid-phase peptide synthesis by v-HPLC/fast atom bombardment mass spectrometry. Peptide research 4 40-46, 1991. 

J. L. Aubaganac, C. Enjalbal, et al.. Application of time-of-flight secondary ion mass spectrometry to in situ monitoring of solid-phase peptide synthesis on the multipin system, J. Mass Spectrom. 33, 1094-1103 (1998). 

The techniques in solid-phase organic chemistry complement those from solid-phase peptide assembly. Synthesis is performed on resins using anchoring linkages and orthogonal protection schemes. Molecules are cleaved from the solid support, purified (see Chapter 18), and then analyzed by high-performance liquid chromatography or mass spectrometry (see Chapters 19 and 20). Reactions can be performed in a simple apparatus that... 

The assembly of the p- and y-amino-acid building blocks to peptidic chains was achieved by simply using the established methods of peptide synthesis - in solution [6], on solid phase [11], or in a synthesizer machine [39] also, the so-called native ligation can be applied with p-peptides [54]. Furthermore, the methods of analyzing and studying the structures of a-peptides and natural proteins can mostiy be applied to P-peptides as well (the same is true for y-peptides [51,55-60]). These methods are CD [35,37] and NMR [6, 49] spectroscopy, mass spectrometry [27,35], X-ray analysis [6,21,24,25,36], molecular dynamics (MD) calculations [9,13,18,31,38] and biological investigations [6, 15,20,26,30,41-43,45,46,48]. All of this sounds like routine, but the results are rather spectacular. 

Truncated sequences, core sequences, incompletely synthesized peptides. Imperfect conversion during acylation or deprotection of the temporary protecting group in solid-phase synthesis may lead to mismatch sequences that lack some amino acids and truncated sequences (core sequences). They occur, when the accessibility or reactivity of the peptide bound to the solid phase is insufficient difficult sequences). Truncated sequences may be classically identified and quantified by a modified Edman degradation, also called preview analysis. Alternatively, mass spectrometry provides efficient tools for the identification of truncated sequences. 

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